Discrimination of non-infectious SARS-CoV-2 particles from fomites by viability RT-qPCR

Cuevas-Ferrando, Enric; Girón-Guzmán, Inés; Falcó, Irene; Pérez-Cataluña, Alba; Díaz-Reolid, Azahara; Aznar, Rosa; Randazzo, Walter; Sánchez, Gloria

The ongoing coronavirus 2019 (COVID-19) pandemic constitutes a concerning global threat to public health and economy. In the midst of this pandemic scenario, the role of environment-to-human COVID-19 spread is still a matter of debate because mixed results have been reported concerning SARS-CoV-2 stability on high-touch surfaces in real-life scenarios. Up to now, no alternative and accessible procedures for cell culture have been applied to evaluate SARS-CoV-2 infectivity on fomites. Several strategies based on viral capsid integrity have latterly been developed using viability markers to selectively remove false-positive qPCR signals resulting from free nucleic acids and damaged viruses. These have finally allowed an estimation of viral infectivity. The present study aims to provide a rapid molecular-based protocol for detection and quantification of viable SARS-CoV-2 from fomites based on the discrimination of non-infectious SARS-CoV-2 particles by platinum chloride (IV) (PtCl(4)) viability RT-qPCR. An initial assessment compared two different swabbing procedures to recover inactivated SARS-CoV-2 particles from fomites coupled with two RNA extraction methods. Procedures were validated with human (E229) and porcine (PEDV) coronavirus surrogates, and compared with inactivated SARS-CoV-2 suspensions on glass, steel and plastic surfaces. The viability RT-qPCR efficiently removed the PCR amplification signals from heat and gamma-irradiated inactivated SARS-CoV-2 suspensions that had been collected from specified surfaces. This study proposes a rapid viability RT-qPCR that discriminates non-infectious SARS-CoV-2 particles on surfaces thus helping researchers to better understand the risk of contracting COVID-19 through contact with fomites and to develop more efficient epidemiological measures.